NG2 cell dynamics in the normal and injured adult central nervous system

描述（由申请人提供）：哺乳动物CNS含有一组不同的神经胶质细胞，其支持神经元、调节神经元活性并使损伤后能够修复。除了星形胶质细胞、少突胶质细胞和小胶质细胞外，现在认识到CNS还含有第四类表现出独特性质的胶质细胞。这些丰富的神经胶质细胞表达血小板衍生生长因子（PDGF 1 R）的α受体和硫酸软骨素蛋白聚糖NG 2，并被称为“NG 2+神经胶质细胞（NG 2细胞）"。NG 2细胞延伸高度分支的突起，装饰有小的丝状伪足样突起，并且以高度有序或“平铺”的方式均匀分布在成年CNS的整个灰质和白色物质区域，类似于星形胶质细胞和小胶质细胞。由于这些细胞作为少突胶质细胞的祖细胞，NG 2细胞的组织和持续动力学可能对于替代通过正常衰老或在诸如多发性硬化症的疾病中急性脱髓鞘而丢失的少突胶质细胞至关重要。然而，NG 2细胞的细胞动力学尚未在哺乳动物CNS中进行体内研究，并且对调节这些细胞在成年CNS中的形态、分布和增殖的因子知之甚少。为了监测NG 2细胞在成年大脑中的动态行为，我们最近开发了转基因小鼠（NG 2-lck-EGFP小鼠），其允许在体内可视化NG 2细胞。使用这些小鼠，可以在静息状态或损伤后观察到活体小鼠皮质中的NG 2细胞，这表明这种方法可以用于帮助研究成年大脑中调节NG 2细胞过程动力学，增殖和分化的机制。在这里，我建议使用NG 2细胞在体感皮层的双光子成像研究成年小鼠大脑内的NG 2细胞的行为。我将在数小时、数天和数周的时间内进行延时成像，以确定NG 2细胞过程的结构动力学，并确定细胞间接触在调节其形态和增殖中的作用。NG 2细胞表达离子型谷氨酸受体，并且在神经胶质细胞中是独特的，因为它们与神经元形成直接突触，提供了神经元可以调节其行为的手段。为了评估NMDA受体信号传导是否调节NG 2细胞的动态行为及其对局灶性损伤的反应，我将在转基因小鼠中进行类似的实验，其中NMDA受体已从NG 2细胞中选择性删除。通过监测NG 2细胞在正常和受损大脑中的体内动态行为，这些研究将进一步加深我们对指导这些无处不在的祖细胞行为的机制的理解。由于这些细胞有能力取代少突胶质细胞，并有助于形成胶质瘢痕，这些研究可能有助于确定加速少突胶质细胞再生和髓鞘修复的新方法，以及增强从各种CNS损伤中恢复的新策略。 公共卫生相关性：该提案旨在了解调节成人大脑中NG 2细胞动态行为的机制。由于NG 2细胞具有替代少突胶质细胞并促进胶质瘢痕形成的能力，从这些研究中获得的信息可能导致开发新的策略，以促进多发性硬化症等疾病中髓鞘的快速修复，并加速CNS损伤（如中风）后的恢复。

DESCRIPTION (provided by applicant): The mammalian CNS contains a diverse group of glial cells that support neurons, modulate neuronal activity, and enable repair following injury. In addition to astrocytes, oligodendrocytes and microglial cells, it is now recognized that the CNS contains a fourth class of glial cell that exhibits unique properties. These abundant glial cells express the alpha receptor for platelet derived growth factor (PDGF1R), and the chondroitin sulfate proteoglycan NG2, and have been termed "NG2+ glial cells (NG2 cells)". NG2 cells extend highly branched processes decorated with small filopodia-like protrusions, and are evenly distributed throughout gray and white matter regions of the adult CNS in a highly ordered or "tiled" manner, similar to astrocytes and microglial cells. As these cells serve as progenitors to oligodendrocytes, the organization and ongoing dynamics of NG2 cells may be crucial for replacing oligodendrocytes lost through normal aging or following acute demyelination in diseases such as multiple sclerosis. However, the cellular dynamics of NG2 cells have not been studied in the mammalian CNS in vivo, and little is known about the factors that regulate their morphology, distribution, and proliferation of these cells in the adult CNS. To monitor the dynamic behavior of NG2 cells in the adult brain, we recently developed transgenic mice (NG2-lck-EGFP mice) that allow the visualization of NG2 cells in vivo. Using these mice, NG2 cells in the cortex of living mice can be observed during resting-states or after injury, indicating that this approach can be used to help investigate the mechanisms regulating NG2 cell process dynamics, proliferation, and differentiation in the adult brain. Here, I propose to use two-photon imaging of NG2 cells in the somatosensory cortex to study the behavior of NG2 cells within the adult mouse brain. I will perform time-lapse imaging over a period of hours, days and weeks to define the structural dynamics of NG2 cell processes, and to determine the role of cell-cell contact in the regulation of their morphology and proliferation. NG2 cells express ionotropic glutamate receptors, and are unique among glial cells in that they form direct synapses with neurons, providing a means by which neurons could regulate their behavior. To evaluate whether NMDA receptor signaling regulates the dynamic behavior of NG2 cells and their response to focal injury, I will perform similar experiments in transgenic mice in which NMDA receptors have been selectively deleted from NG2 cells. By monitoring the dynamic behavior of NG2 cells in vivo in the normal and injured brain, these studies will further our understanding of the mechanisms that guide the behavior of these ubiquitous progenitors. As these cells have the capacity to replace oligodendrocytes and contribute to the formation of glial scars, these studies may help identify new approaches for accelerating oligodendrocyte regeneration and myelin repair, as well as new strategies for enhancing recovery from diverse CNS injuries. PUBLIC HEALTH RELEVANCE: This proposal seeks to understand the mechanisms that regulate the dynamic behavior of NG2 cells in the adult brain. As NG2 cells have the ability to replace oligodendrocytes and contribute to the formation of glial scars, information gained from these studies may lead to the development of new strategies to promote rapid repair of myelin sheaths in diseases such as multiple sclerosis, and accelerate recovery following CNS injuries such as stroke.